Physical adsorption of lipase onto mesoporous silica

Lipases are versatile enzymes that catalyze the hydrolysis of the ester bonds of lipids. For this study commercial enzyme Lipolase 100L was studied as a source of lipase. Two silica preparations, varying in mesh size (Silica70-230 and Silica230-400), were used for immobilisation of Lipolase 100L (1:10 = lipase: tris buffer). It was observed that activity assay gave the highest values when treated with 2% of guteraldehyde. 9 mM p- NPP gave the best results as suitable substrate. The enthalpy of activation for silica immobilized lipase decreased with an increase in temperature whereas Km and Vmax was analysed on the Lineweaver Burk Plot, the values were observed for S70-230 lipolase (Km = 66.67 mM ,Vmax = 3.34 U/mg/min., kcat =35.22 s-1 and specificity constant =10.56 s-1Mm-1 ), S230-400 lipolase (Km = 52.64 mM , of Vmax is 2.32 U/mg/min., kcat =27.80 s-1 and specificity constant =12.00 s-1Mm-1 ) and pure lipolase enzyme (Km value as 62.5mM , Vmax 2.1875 U/mg/min , kcat =33.02 s-1 and specificity constant =15.09 s-1Mm-1 ).The thermostability of silica immobilized showed that the half-life (t1/2) of immobilized lipase was approximately 30 minutes at 550C. Among chelating and denaturing agents like SDS, EDTA, mercaptoethanol and PEG affected the activity by lowering its value too. The lipase immobilizes silica is reusable upto 5 cycles during which it retained it activity upto 50%.